Hi, I'm trying to set up a contrast matrix for my microarray experiment and I'm not sure whether I did it in the right way. The experiment has a 2x2 factorial (S or T; c or d) design and samples (Sc, Sd, Tc, Td) were hybridized in a loop design. I want to include the dye effect in the design and calculate the contrasts for d vs. c; dT-cT; dS-cS and the interaction [(dT-cT)-(dS-cS)]. Following the example for a single factor exp. in the user guide with dye effect I tried this: > design <- modelMatrix(targets, ref="Sc") Found unique target names: Sc Sd Tc Td > design <- cbind(Dye=1, design) > design Dye Sd Tc Td Slide03 1 1 0 0 Slide04 1 0 1 -1 Slide05 1 -1 1 0 Slide06 1 0 0 1 Slide09 1 0 -1 0 Slide10 1 1 0 -1 Slide19 1 -1 1 0 Slide20 1 1 0 0 Slide21 1 0 -1 1 Slide22 1 0 0 -1 Slide23 1 -1 0 0 Slide25 1 0 -1 1 Slide27 1 -1 0 0 Slide28 1 0 -1 1 > contrast.matrix <- cbind( + "d-c"=c(0,0.5,-0.5,0.5), + "dT-cT"=c(0,0,-1,1), + "dS-cS"=c(0,1,0,0), + "Interaction"=c(0,-1,-1,1)) > rownames(contrast.matrix)<-colnames(design) > contrast.matrix d-c dT-cT dS-cS Interaction Dye 0.0 0 0 0 Sd 0.5 0 1 -1 Tc -0.5 -1 0 -1 Td 0.5 1 0 1 Is this correct? Thanks in advance, Thomas Degenkolbe